蒜芥茄黄萎病抗性相关基因SsWRKY22和SsWRKY33的克隆与表达分析  

Cloning and Expression Analysis of Solanum sisymbriifolium Verticillium Wilt Resistance-Related Genes SsWRKY22 and SsWRKY33

作　　者：尹梦莹 蔚亚楠 龚亚菊[2] 桂敏[2] 鲍锐[2] 黎志彬[2] 吴丽艳[2] 杜光辉[1] Yin Mengying;Yu Yanan;Gong Yaju;Gui Min;Bao Rui;Li Zhibin;Wu Liyan;Du Guanghui(Institute of Resource Plants,Yunnan University,Kunming,650500;Horticultural Research Institute Yunnan Academy of Agricultural Sciences,Kunming,650205)

机构地区：[1]云南大学资源植物研究院,昆明650500 [2]云南省农业科学院园艺作物研究所,昆明650205

出　　处：《分子植物育种》2024年第8期2464-2472,共9页Molecular Plant Breeding

基　　金：国家自然科学基金地区基金项目(31960594);云南省农业基础研究联合专项面上项目(2018FG001-022);云南省应用基础研究面上项目(2019FB059)共同资助。

摘　　要：黄萎病是茄子生产中的重要病害之一,严重影响茄子的产量和品质。WRKY转录因子在植物抗病中扮演着重要的角色,但在茄子黄萎病抗性中的作用不明。本研究从黄萎病高抗材料野生蒜芥茄的转录组数据中,筛选获得WRKY家族中的SsWRKY22和SsWRKY33基因,利用PCR技术克隆获得目的基因,并对其进行生物信息学和表达模式分析。结果表明,SsWRKY22和SsWRKY33基因的ORF序列大小分别为528和1073 bp,编码175个和357个氨基酸;蛋白分子量分别为19.19和40.46 kD,均属于亲水性不稳定蛋白,其中,Ss WRKY22编码的蛋白属于酸性蛋白,而Ss WRKY33编码的蛋白属于碱性蛋白,亚细胞定位预测两个基因编码的蛋白均定位于细胞核上;此外,SsWRKY22和SsWRKY33分别属于Ⅱ类和Ⅰ类WRKY转录因子,具有不同的蛋白结构。系统发育分析结果表明,Ss WRKY22与野生潘那利番茄、番茄和马铃薯中的WRKY22亲缘关系较近;而Ss WRKY33与辣椒中的WRKY33亲缘关系最近。利用RT-qPCR对SsWRKY22和SsWRKY33基因在蒜芥茄不同部位(根,茎,叶)的表达模式进行分析,发现二者的表达具有组织特异性,且在根部的表达量均为最高。在接种黄萎病病菌后,SsWRKY22和SsWRKY33基因在蒜芥茄根部的表达量显著下调,表明SsWRKY22和SsWRKY33可能是蒜芥茄响应黄萎病胁迫的负调控因子。本研究首次克隆了云南野生蒜芥茄的WRKY转录因子中的SsWRKY22和SsWRKY33基因并解析其表达模式,为后续SsWRKY22和SsWRKY33的功能鉴定提供基础。Verticillium wilt is a major disease that seriously affects eggplant yield and quality.WRKY transcription factors play a key role in plant resistance,while the knowledge about the resistance to eggplant verticillium wilt are deficient.In this study,two WRKY genes,named SsWRKY22 and SsWRKY33,were screened from the transcriptome database of Solanum sisymbriifolium which is highly resistant to verticillium wilt.The target genes were cloned for bioinformatics analysis.Then the expression pattern of two genes were analyzed by RT-qPCR.The results showed that the full length ORF of SsWRKY22 and SsWRKY33 are 528 and 1 073 bp,encoding 175 and 357amino acid,respectively.The molecular weights of two proteins are 19.19 and 40.46 kD,respectively,which belong to hydrophilic unstable proteins.The Ss WRKY22 protein is acidic protein,while the Ss WRKY33 protein is basic protein.Subcellular localization predicted that both proteins are located in the nucleus.In addition,Ss WRKY22 and Ss WRKY33 with different protein structure belong to class Ⅱ and class Ⅰ WRKY transcription factors,respectively.Phylogenetic analysis showed that Ss WRKY22 is closely related to WRKY22 in Solanum pennellii,S.lycopersicum and S.tuberosum,while the SsWRKY33 is the most closely related to WRKY33 in Capsicum annuum.The RT-qPCR analysis indicated that the expression levels of SsWRKY22 and SsWRKY33 are tissue-specific and the highest in roots.After inoculation by Verticillium dahliae,the expression levels of SsWRKY22 and Ss WRKY33were significantly down-regulated in the root,suggesting that both genes may be negative regulatory factors in response to verticillium wilt stress.In this study,WRKY transcription factor genes were firstly cloned and their expression patterns were also analyzed in S.sisymbriifolium,which will lay a foundation for the follow-up functional characterization of SsWRKY22 and SsWRKY33.

关 键 词：蒜芥茄 黄萎病 SsWRKY22 SsWRKY33 表达模式 

分 类 号：S436.411[农业科学—农业昆虫与害虫防治]